Chloramphenicol is a well known broad spectrum antibiotic. Unfortunately it produces aplastic anemia in one out of 20,000 people who are treated. This laboratory has reported earlier that chloramphenicol is metabolically activated to some "reactive intermediate(s)" which bind covalently to tissue macromolecules followed by alteration in cellular integrity and probably cell death. In order to understand the mechanism of the formation of reactive intermediate(s) of chloramphenicol protein containing covalently bound chloramphenicol (14C or 3H) was hydrolyzed with Pronase and the hydrolyzates analyzed in an Amino Acid Analyzer. The results obtained so far indicate that most likely bioactivated chloramphenicol binds covalently to four amino acids in the protein of liver microsomes.